The ultimate goal of systems neuroscience is to explain human behavior, but as the human nervous system contains billions of neurons, simpler but still relevant model systems may assist in approaching this goal. To gain insight into simple neuronal circuits with relevance in humans, temperature sensing by Drosophila larvae serves as a meaningful but scaled down model due to broad conservation of sensory TRP (transient receptor protein) superfamily cation channels. Although a consensus model exists for the larval warm circuit, the cold-sensing circuit remains controversial, with disagreement over the role of relevant temperature-sensing molecules, such as TRPL (TRP-like), a TRP channel famous for its canonical role in phototransduction for fly vision. Importantly, TRP channel defects are relevant for human diseases and impairments, such as deafness, vestibular difficulties, neurological disorders, cardiac hypertrophy, gastroesophageal reflux disease, bladder diseases, several cancers, and chronic obstructive pulmonary disease. In addition, the location of larval cold-sensing neurons has not been firmly established. Here, Drosophila larvae are used to study thermotaxis, or navigation in response to changes in environmental temperature, to provide insights into decision-making behaviors in higher organisms. This proposal focuses on the cold-sensing circuit to characterize specific cold-responsive neurons and to quantitatively examine the cold sensor candidate TRPL. To accomplish these aims, genetic tools, such as genetically encoded calcium indicators, optogenetics, and the GAL4-UAS system, are used in conjunction with custom hardware and software designed to quantitatively track thermotaxis and perform three-dimensional in vivo calcium imaging in awake immobilized larvae subjected to temperature modulations. PUBLIC HEALTH RELEVANCE: Difficulties in accurately sensing one's environment lead to a wide variety of diseases and impairments, including deafness, vestibular difficulties, neurological disorders, cardiac hypertrophy, gastroesophageal reflux disease, bladder diseases, several cancers, and chronic obstructive pulmonary disease. Because the same conserved molecules are used for sensing in humans as well as in Drosophila (fruit fly) larvae, which are much simpler and easier to study, probing the mechanisms behind larval temperature sensing is relevant for public health. The goal of this proposal is to gain an understanding of the larval cold-sensing circuit as a relevant analogy in humans.